Improved test apparatus has been provided for detection of substance for agglutination reactions. As examples of such improved test apparatus there can be mentioned U.S. Pat. Nos. 4,596,695; 4,597,944 and 4,775,515 of Hugh V. Cottingham and the agglutination slide device disclosed in U.S. Pat. No. 5,019,351 of Peter Schulz.
The agglutination test is based on latex agglutination-inhibition principles in which there is competition for binding to an antibody between the analyte and latex particles coated with an analyte analog or conjugate of the analyte. A sample is placed in the mixing well of a slide apparatus of the type disclosed in the aforementioned U.S. Pat. No. 5,019,351 along with the latex particles coated with the analyte analog or conjugate and the antibody. The mixture traverses the capillary path of the slide apparatus by capillary action to a viewing area. If there is no analyte in the sample, the latex particles with the analyte analog or conjugate form large clumps of particles (agglutinates) by binding to the antibody. However, when analyte is present in the sample, the analyte competes with the labeled latex particles for reaction with the antibody and the analyte preferentially binds to the antibody and inhibits or prevents reaction of the antibody with the labeled latex particles and thereby inhibits or prevents agglutination of the latex particles. Thus, the presence of agglutination of the latex particles is evidence of the absence of the analyte from the sample, whereas the absence of significant agglutination of the latex particles is evidence of the presence of the analyte in the sample, when the presence or absence of agglutination of the latex particles is visually observed in the viewing area of the slide apparatus.
Using the aforementioned technology and the agglutination slide apparatus disclosed in the aforesaid patents, test kits have been marketed for the easy, rapid determination of various biological substances such as hormones, tumor markers and the like and also for drugs of abuse, such as amphetamines, barbituates, cocaine, marijuana, morphines, phencyclidine and the like from biological samples, such as urine, blood or other body fluids. Such test apparatus and assay procedure have permitted easy and rapid field assays of biological fluids for such biological substances and drugs of abuse. Such rapid assays are able to be readily conducted in the field and require no instrumentation for analysis of the results. A visual qualitative result is observable in the slide viewing area, generally within about three to five minutes or less from the time of mixing the sample, labeled latex particles and antibody in the mixing well of the slide apparatus.
The use of such agglutination reaction slide apparatus has been highly beneficial, permitting on-site field analysis, that is, in a non-laboratory setting, by a simple, easy procedure without requiring other instrumentation. An example of such an assay test system for drugs of abuse is the ONTRAK test system sold by Roche Diagnostic Systems, Inc. of Somerville, N.J. Thus, this technology has replaced, at least in part, screening assay procedures previously required to be used, such as thin layer chromatography or liquid chromatography, enzyme immunoassay, fluorescence polarization immunoassay or radioimmuno-assay, requiring some type of instrumentation.
However, such agglutination reaction slide assay technology has been found to have an especially limiting drawback, namely that the concentration of analyte in the sample being analyzed must be at a relatively high concentration level of at least about 50 ppb or more in order to produce agglutination-inhibition to provide the desired visual result since generally only about a 11 .mu.l sample volume in a total reaction volume of about 160 .mu.l is able to be put into the mixing well of a slide apparatus. This drawback has prevented such agglutination slide reaction assay technology from being usable to detect analytes such as pesticides or environmental toxins in environmental samples, such as water, effluent water, soil, sludge, manure wastes or sediments or the like, where the pesticide residues or environmental toxins are or may be present only in very low concentration, such as about 1 ppb or less.
Similarly, because of the generally low levels of LSD, i.e. lysergic acid diethylamide, in biological samples, e.g. below about 1.5 ppb in urine, it has not been possible to utilize the ONTRAK test system for detecting this common drug of abuse.
It has also previously been proposed to detect analytes by use of affinity chromatography techniques where an affinity membrane with a functional group and antibody attached directly to the membrane was permitted to come into intimate contact with the sample suspected of containing the analyte of interest. However, there is insufficient binding capacity for a number of reasons, including low surface binding area and loss of functionality of the antibody, and therefore generally only about 20% to about 40% of analyte is able to be recovered for detection and assay. Thus, such affinity chromatography techniques have also not provided a satisfactory field assay procedure for detection of 10 ppb or 1 ppb levels of pesticides and environmental toxins in environmental samples.
It is therefore highly desirable that an agglutination slide reaction assay system be available for quick, easy, in-the-field assays samples containing low level of analytes such as pesticides or environmental toxins in environmental samples and low levels LSD or other drugs of abuse in biological samples.